TW202337437A - Aqueous pharmaceutical preparations for injection, dry pharmaceutical preparations, kits and preparation methods thereof - Google Patents

Abstract

The present invention relates to a dry pharmaceutical formulation obtainable by drying an aqueous pharmaceutical formulation according to the present invention. Provide pharmaceutical formulations comprising high concentrations of dissolved QTX125 at physiological pH which are stable and have low toxicity. The present invention relates to an aqueous pharmaceutical formulation for injection. The aqueous pharmaceutical formulations according to the present invention mean that the compound QTX125 can be dissolved at a concentration of 7.5 mg/ml or more in a formulation with a pH of between pH7 and pH8, which is suitable for injection. The aqueous pharmaceutical formulations according to the present invention are non-toxic in mammals at dosages of up to 200 mg/kg and hence are suitable for use in methods of treatment in mammals, such as methods of treating cancer.

Description

Aqueous pharmaceutical preparations for injection, dry pharmaceutical preparations, kits and their preparation methods and uses

The present invention relates generally to pharmaceutical formulations and the field of pharmacy, and specifically to pharmaceutical formulations comprising certain histone deacetylase inhibitors and cyclodextrins or derivatives thereof. The invention also relates to the use of such preparations, for example, for the treatment of cancer or autoimmune diseases.

Histone deacetylase (HDAC)

Histone deacetylase (HDAC) constitutes an interesting therapeutic target for the treatment of cancer (see P.A. Marks et al. Nature Rev. Cancer 2001, 1, 194; J. E. Bolden et al. Nature Rev. Drug Discov. 2006, 5, 769; P. Gallinari et al. Cell Res. 2007, 17, 195; K. B. Glaser Biochem. Pharmacol. 2007, 74, 659; L. Panetal. Cell. Mol. Immunol. 2007, 4, 337; M . Haberland et al. Nature Rev. Genetics 2009, 10, 32; Y. Zhang et al. Curr. Med. Chem. 2008, 15, 2840; S. Ropero, M. Esteller Mol. Oncol. 2007, 1, 19) and other diseases, such as those related to the central nervous system, such as autoimmune diseases (see A.G. Kazantsev, L.M. Thompson Nature Rev. Drug Discov. 2006, 7, 854).

Several families of HDAC inhibitors (HDACis) have been designed and their general structures can be found in different reviews (see A. Villar-Garea, M. Esteller Int. J. Cancer 2004, 112, 171; T.A. Miller et al. J. Med. Chem. 2003, 46, 5097; T. Suzuki, N. Miyata Curr. Med. Chem. 2005, 12, 2867; M. Paris et al. J. Med. Chem. 2008, 51, 1505). The general structure of these inhibitors consists of a cyclic structure, a spacer, and the ability to bind to class I (HDAC1, HDAC2, HDAC3, and HDAC8), class II (HDAC4, HDAC5, HDAC6, HDAC7, HDAC9, and HDAC10) and class IV (HDAC11 ) The active center of different HDAC isomers consists of Zn(II) cation-bound chelating groups.

The mechanism of action of HDAC inhibitors can be explained by their antagonistic properties against histone deacetylase, which is involved in the regulation of apoptosis, cell growth, tumor progression, cancer metastasis, cell adhesion, etc. process. These properties prevent the binding of HDACs to their natural ligands, which can be histones or cytoplasmic proteins such as tubulin, and their normal catalytic initiation, i.e., the ε-N-acetyllysine residue present in these proteins. Deacetylation.

Despite similar inhibition patterns, some selectivity for inhibition of different HDAC isoforms was occasionally observed (see J.C. Wong et al. J. Am. Chem. Soc. 2003, 125, 5586; G. Estiu et al. J. Med. Chem. 2008, 51, 2898). The selectivity mentioned has therapeutic implications (see K. V. Butler, A. P. KozikowskiCurr. Pharm. Design 2008, 14, 505; T. C. Karagiannis, A. El-OstaLeukemia 2007, 21, 61).

HDAC inhibitors

An important class of HDAC inhibitors are trisubstituted pyrrole derivatives linked via aromatic and heteroaromatic groups to a chelating group, as described for example in WO2011/039353. These compounds have been shown to be effective in treating cancer (see WO2011/039353).

Additionally, these compounds have been shown to be effective in treating several autoimmune diseases. For example, these compounds have been shown to be effective in animal models of autoimmune hepatitis and autoimmune encephalomyelitis (see WO 2018/087082).

One particularly promising compound is 3-(3-furyl)-N-{4-[(hydroxylamino)carbonyl]benzyl}-5-(4-hydroxyphenyl)-1pyrrole-2-methamide (Referred to as QTX125 in this article). QTX125

QTX125 is a highly selective and potent HDAC 6 inhibitor. It has shown high anti-tumor efficacy in mantle cell lymphoma (see Perez-Salvia, M. et al haematologica 2018;103:e540), lung and pancreatic cancer xenograft mouse models. QTX125 also showed high efficacy in two different mouse models of multiple sclerosis (see WO2018/087082).

However, hydroxamic acids such as QTX125 are known to have very low solubility in water (see Patre, S. et al. 2011 International Conference on Environment and BioScience IPCBEE vol. 21 (2011)) and in order to dissolve QTX125 in aqueous solutions it is often necessary to use High pH. QTX125 also exhibits physical and chemical instability in solution.

For injectable formulations of QTX125, it is desirable to have a solubility of 7.5 mg/mL or greater in formulations at physiological pH (i.e., from pH 7 to pH 8). This is particularly important from the perspective of avoiding pain during injection, since compositions with particularly high or low pH tend to be more painful. Without being bound by any theory, it is believed that formulations for infusion administration should be close to the physiological pH range (pH 7-8) and that for large dose administration the pH of buffered solutions should be 4-8 and the pH of unbuffered solutions should be 3 -9. Injectable formulations should also remain stable for an extended period of time between preparation and administration and have low toxicity.

Therefore, there remains a need in the art to provide pharmaceutical formulations that include high concentrations of dissolved QTX125 at physiological pH and are stable and have low toxicity.

In order to more fully describe and disclose the present invention and the prior art to which this invention pertains, numerous patents and publications are cited herein. Full citations for these references are provided in this article. Each of these references is incorporated by reference into this disclosure in its entirety.

The present inventors have developed a pharmaceutical formulation that helps solve the above practical problems.

Accordingly, in a first aspect, the present invention provides an injectable pharmaceutical preparation, comprising:

Compounds of formula I, Formula I

and compounds of formula II, Formula II

where each ^R1 is independently selected from -H or group;

Wherein, R ² is absent or is C _1-4 alkyl;

Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and

R ³ is independently selected from -H or C _1-4 alkyl;

Wherein, the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2; and

Wherein, the pH of the pharmaceutical preparation is between pH 7 and pH 8.

Preferably, the present invention relates to an aqueous pharmaceutical preparation for injection comprising a compound of formula I, Formula I

and sulfobutyl ether-β-cyclodextrin (SBβCD),

Wherein, the molar ratio of the compound of formula I to sulfobutyl ether-β-cyclodextrin is 1:50-1:2; and

Wherein, the pH of the pharmaceutical preparation is between pH 7 and pH 8.

In a second aspect, the invention relates to an aqueous pharmaceutical preparation for injection, comprising:

Compounds of formula I, Formula I

and compounds of formula II, Formula II

where each ^R1 is independently selected from -H or group;

Wherein, R ² is absent or is C _1-4 alkyl;

Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and

R ³ is independently selected from -H or C _1-4 alkyl;

Wherein, the pharmaceutical preparation can be obtained by methods including:

Step 1: Prepare a mixture including water, a compound of formula I and a compound of formula II, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2; and

Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8.

In another aspect, the invention relates to a dry pharmaceutical preparation obtainable by drying an aqueous pharmaceutical preparation according to the invention, preferably obtainable by freeze-drying an aqueous pharmaceutical preparation according to the invention.

In another aspect, the invention relates to a kit comprising said dry pharmaceutical formulation and a pharmaceutically acceptable grade of water, buffer solution or saline solution for reconstituting the dosage form.

In another aspect, the invention relates to an aqueous pharmaceutical formulation or a dry pharmaceutical formulation as described herein for use in the manufacture of a medicament.

In another aspect, the invention relates to the use of an aqueous pharmaceutical formulation or a dry pharmaceutical formulation as described herein for the preparation of a medicament for the treatment of cancer or autoimmune diseases.

In another aspect, the invention relates to the use of an aqueous pharmaceutical formulation as described herein, or a liquid pharmaceutical formulation obtainable by reconstituting a dry pharmaceutical formulation, as a medicament.

In another aspect, the invention relates to the use of an aqueous pharmaceutical formulation as described herein or a liquid pharmaceutical formulation obtainable by reconstituting the formulation for the treatment of cancer or autoimmune diseases.

In another aspect, the invention relates to a method of treatment comprising administering to a patient an aqueous pharmaceutical formulation as described herein or a liquid pharmaceutical formulation obtainable by reconstituting a dry pharmaceutical formulation.

In another aspect, the invention relates to a method of treating cancer or an autoimmune disease, comprising administering to a patient an aqueous pharmaceutical formulation or a liquid pharmaceutical formulation obtainable by reconstituting a dry pharmaceutical formulation as described herein.

As those skilled in the art will appreciate, features and preferred embodiments of one aspect of the invention will also apply to other aspects of the invention.

definition

Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. Methods and materials for use in the present disclosure are described herein; other suitable methods and materials known in the art may also be used. The materials, methods, and examples are illustrative only and not restrictive. All publications, patent applications, patents, sequences, database entries, and other references mentioned herein are incorporated by reference in their entirety. In case of conflict, the present specification, including definitions, will control.

The term "about" before a stated value means that the value may have an uncertainty of ±20%, preferably ±10%, ±5%, ±2%, ±1% of the stated value.

The term "C ₁ -C ₄ alkyl" refers to a straight or branched hydrocarbon chain consisting of carbon and hydrogen atoms, containing no unsaturation, and having 1 to 4 carbon atoms, preferably 1 to 3 carbon atoms (" C ₁ -C ₃ alkyl") and which is attached to the rest of the molecule by a single bond, such as and without limitation methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl Key et al.

The term "room temperature" refers to the ambient temperature of a typical laboratory, usually between 20°C and 30°C, preferably around 25°C at atmospheric pressure.

The term "dry state" refers to a formulation that has been dried. Optionally, a dry formulation may refer to a solid material having a residual water content of less than 10%, preferably less than 8%, preferably less than 5%, preferably from about 0.1% to about 5%. The residual water content can be determined using Karl Fischer titration.

The term "lyophilized" or "freeze-dried" refers to substances that have been obtained and/or are obtainable by freeze-drying liquid preparations, resulting from a drying process in which the material to be dried is first frozen and then the ice is removed by vacuum sublimation or Freezing solvent.

The term "reconstituted" refers to the dry (lyophilized) formulation with a pharmaceutically acceptable liquid, such as a pharmaceutically acceptable grade of water (preferably sterile), a pharmaceutically acceptable buffer solution, or a saline solution, To convert a powder (or solid compound) into a suspension or solution that can be administered to a patient by injection. The term "reconstituted dry pharmaceutical preparation" relates to a liquid pharmaceutical preparation obtainable by reconstituting a dry pharmaceutical preparation according to the invention in a pharmaceutically acceptable liquid.

The term "injection" refers to any form of injection known to those skilled in the art, such as subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital , intraperitoneal, intratracheal, subepidermal, intraarticular, subarachnoid space and intrasternal. Injection can refer to infusion procedures (eg, continuous administration) as well as bolus (intermittent) administration.

The term "pharmaceutically acceptable salt" refers to a salt that when administered to a recipient provides (directly or indirectly) a compound as described herein. "Pharmaceutically acceptable" preferably refers to compositions and molecular entities that are physiologically tolerable and generally do not produce allergic reactions or similar adverse reactions, such as stomach problems, dizziness, etc., when administered to humans or animals. Preferably, the term "pharmaceutically acceptable" means that it is approved by a regulatory agency of a state or federal government or is included in the United States Pharmacopeia or other recognized pharmacopeia for use in animals, and more particularly in humans.

The term "treatment" or "treatment" refers to the administration of a compound or pharmaceutical composition of the invention to ameliorate or eliminate a disease or one or more symptoms associated with a disease. The term "prevention" or "prevention" includes reducing the risk of the emergence or progression of disease.

If not stated otherwise, "%" refers to weight %.

The present invention has many advantageous features, including those listed below.

First, the aqueous pharmaceutical preparation according to the present invention means that the compound QTX125 can be dissolved in a preparation suitable for injection between pH 7 and pH 8 at a concentration of 7.5 mg/mL or higher.

Secondly, the aqueous pharmaceutical formulation according to the present invention is non-toxic to mammals at doses up to 200 mg/kg and is therefore suitable for use in methods of treating mammals, such as methods of treating cancer.

Thirdly, aqueous pharmaceutical formulations according to the present invention are stable over a longer period of time, for example up to 3 months at 5°C. Aqueous pharmaceutical formulations can also be freeze-dried without resulting in any loss of activity of the formulation upon reconstitution.

Fourth, dry (lyophilized) pharmaceutical formulations according to the present invention exhibit high stability at room temperature for a period of at least 3 months, which may eliminate the need for cold chain transportation of these formulations. These formulations are stable and maintain a pH between pH 7 and pH 8 once reconstituted even after long storage.

Compounds of formula I

One aspect of the present invention focuses on providing an aqueous pharmaceutical formulation having a compound of formula I (QTX125) dissolved in a high concentration.

The aqueous pharmaceutical preparation includes:

Compounds of formula I, Formula I

and compounds of formula II, Formula II

where each ^R1 is independently selected from -H or group;

wherein R ² is absent or is C _1-4 alkyl;

Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and

R ³ is independently selected from -H or C _1-4 alkyl;

wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2; and

The pH of the above pharmaceutical preparation is between pH 7 and pH 8.

Preferably, the concentration of the compound of formula I dissolved in the aqueous pharmaceutical formulation is 7.5 mg/mL or higher, more preferably 8 mg/mL or higher, more preferably 8.5 mg/mL or higher, more preferably 9 mg/mL. mL or higher, most preferably 9.5 mg/mL or higher.

The concentration of QTX125 dissolved in the formulation can be determined using HPLC as described in the experimental section below.

The maximum concentration at which QTX125 can be dissolved is limited only by the maximum solubility value that QTX125 can achieve. Optionally, the maximum concentration of QTX125 in solution is 50 mg/mL, or 20 mg/mL, or 15 mg/mL.

The compounds of formula I can be incorporated into aqueous pharmaceutical preparations in the form of salts (preferably as pharmaceutically acceptable salts), as solvates, as free salts, as neutral compounds or as prodrugs.

The preparation of salts can be accomplished by methods known in the art. For example, pharmaceutically acceptable salts can be synthesized by conventional chemical methods from the original compound containing a basic residue. Typically, such salts are prepared, for example, by reacting the free salt form of the compound with a suitable salt or acid in water or in an organic solvent or in a mixture of both. In general, non-aqueous media such as diethyl ether, ethyl acetate, ethanol, isopropanol or acetonitrile are preferred. Examples of acid addition salts include inorganic acid addition salts, such as hydrochlorides, hydrobromides, hydroiodates, sulfates, nitrates, phosphates; and organic acid addition salts, such as acetates, horseradish Lenate, fumarate, citrate, oxalate, succinate, tartrate, malate, mandelate, methanesulfonate and p-toluenesulfonate. Examples of addition salts include inorganic salts such as sodium, potassium, calcium, ammonium, magnesium, aluminum and lithium salts, and organic salts such as ethylenediamine, ethanolamine, N,N-dioxane Basic salts of base ethanolamine, triethanolamine, glucosamine and amino acids.

Compounds of formula II

Aqueous pharmaceutical preparations according to the invention further comprise compounds of formula II Formula II

where each ^R1 is independently selected from -H or group;

wherein R ² is absent or is C _1-4 alkyl;

Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and

R ³ is independently selected from -H or C _1-4 alkyl;

The compound of formula II is a β-cyclodextrin backbone, which may be substituted by a number of different functional groups on the hydroxyl group of the β-cyclodextrin (as described above). Cyclodextrins are cyclic oligosaccharides with a ring structure and a hydrophobic/lipophilic central cavity and a hydrophilic outer surface. β-Cyclodextrin is a cyclodextrin composed of 7 glucopyranose units.

Many different types of β-cyclodextrin and their derivatives have been considered safe for use as pharmaceutical excipients. For example, 2-hydroxypropyl-β-cyclodextrin has been shown to be It is well tolerated in a range of animal species (S. Gould et al. Food and Chemical Technology, 43, 1451-1459, 2005) and sulfobutyl ether-β-cyclodextrin (SBβCD) acutely No significant effects were observed in toxicity studies and renal histopathology (Rajewski, RA, 1990. Development and evaluation of the usefulness and parenteral safety of modified cyclodextrins. Lawrence, KS, USA, Univ. Kansas, 251 pp).

Without being bound by any theory, it is believed that cyclodextrins interact with QTX125 compounds to form water-soluble complexes, such as by binding QTX125 molecules into their hydrophobic central cavities.

The compound of formula II is preferably selected from β-cyclodextrin (naturally occurring β-cyclodextrin), (C _1-4 alkyl)-β-cyclodextrin, (C _1-4 alkyl)-β-cyclodextrin Sulfobutyl ether of dextrin, (hydroxy-C _1-4 alkyl)-β-cyclodextrin, C _1-4 alkyl-carboxyalkyl-β-cyclodextrin and C _1-4 alkyl-( Hydroxy-C _1-4 alkyl)-β-cyclodextrin.

More preferably, the compound of formula II is selected from β-cyclodextrin, (C _1-4 alkyl)-β-cyclodextrin, (hydroxy-C _1-4 alkyl)-β-cyclodextrin and (C Sulfobutyl ether of _1-4alkyl )-β-cyclodextrin.

More preferably, the compound of formula II is selected from the group consisting of (hydroxy-C _1-4 alkyl)-β-cyclodextrin and the sulfobutyl ether of (C _1-4 alkyl)-β-cyclodextrin.

More preferably, the compound of formula II is hydroxypropyl-β-cyclodextrin or sulfobutyl ether-β-cyclodextrin (SBβCD).

Most preferably, the compound of formula II is sulfobutyl ether-β-cyclodextrin (SBβCD).

Alternatively, the invention may comprise two or more compounds according to formula II, for example a formulation according to the invention may comprise hydroxypropyl-β-cyclodextrin and sulfobutyl ether-β-cyclodextrin (SBβCD ).

The compounds of formula II can be mixed in the free form or in the form of salts, for example as sodium or potassium salts.

As one skilled in the art will appreciate, the average substitution pattern of cyclodextrins such as SBβCD can vary. SBβCD may have an average of 2 to 8 hydroxyl groups substituted by sulfobutyl ether moieties, preferably 5 to 7 hydroxyl groups substituted by sulfobutyl ether moieties.

The average molecular weight of the compounds of formula II depends on the degree of substitution. The average molecular weight of commercial formulations of sulfobutyl ether-β-cyclodextrin (SBβCD) can vary between 1451–2242 g/mol.

molar ratio of components

In the aqueous pharmaceutical preparation according to the invention, the molar ratio of the compound of formula I to the compound of formula II is from 1:50 to 1:2.

Preferably, the molar ratio of the compound of formula I to the compound of formula II is 1:40 to 1:2, preferably 1:30 to 1:2, preferably 1:25 to 1:2, preferably 1:20 to 1:2 , preferably 1:15 to 1:2, preferably 1:10 to 1:2, preferably 1:9 to 1:2, preferably 1:8 to 1:2, preferably 1:6 to 1:2, more preferably 1: 4.5 to 1:2. Most preferably about 1:2.7.

In certain aspects, the molar ratio of the compound of Formula I to the compound of Formula II is from 1:50 to 1:2.3.

Preferably, the molar ratio of the compound of formula I to the compound of formula II is 1:40 to 1:2.3, preferably 1:30 to 1:2.3, preferably 1:25 to 1:2.3, preferably 1:20 to 1:2.3 , preferably 1:15 to 1:2.3, preferably 1:10 to 1:2.3, preferably 1:9 to 1:2.3, preferably 1:8 to 1:2.3, preferably 1:6 to 1:2.3, more preferably 1: 4.5 to 1:2.3. Most preferably about 1:2.7.

In certain preferred aspects, the molar ratio of the compound of formula I to the compound of formula II is from 1:50 to 1:2.5.

Preferably, the molar ratio of the compound of formula I to the compound of formula II is 1:40 to 1:2.5, preferably 1:30 to 1:2.5, preferably 1:25 to 1:2.5, preferably 1:20 to 1:2.5 , preferably 1:15 to 1:2.5, preferably 1:10 to 1:2.5, preferably 1:9 to 1:2.5, preferably 1:8 to 1:2.5, preferably 1:6 to 1:2.5, more preferably 1: 4.5 to 1:2.5. Most preferably about 1:2.7.

HPLC/MS can be used to determine the concentration of a compound of formula II (eg, sulfobutyl ether-β-cyclodextrin) in solution based on comparison of peak areas with calibration curves of known compounds.

Other additional ingredients

Optionally, the aqueous pharmaceutical formulation additionally includes one or more other pharmaceutically acceptable ingredients known to those skilled in the art, including, but not limited to, pharmaceutically acceptable carriers, diluents, excipients, adjuvants, buffers, pH adjusters, preservatives, antioxidants, bacteriostatic agents, stabilizers, suspending agents, solubilizers, surfactants (e.g., wetting agents), colorants, and isotonic solutes (i.e., that render the preparation compatible with the blood of the intended recipient or other related body fluids isotonic). Suitable carriers, diluents, excipients, etc. can be found in standard pharmaceutical literature. See, for example, Handbook of Pharmaceutical Additives, 2nd ed. (eds. M. Ash and I. Ash), 2001 (Synapse Information Resources, Inc., Endicott, New York, USA), Remington's Pharmaceutical Sciences, 18th ed., Mack Publishing Company , Easton, Pa., 1990; and Handbook of Pharmaceutical Excipients, 2nd edition, 1994.

Optionally, the aqueous pharmaceutical formulation according to the invention further comprises a buffer (ie the formulation further comprises a buffer salt dissolved therein). Optionally, the buffer can be selected from MES, Bis-Tris, ADA, ACES, PIPES, MOPSO, BES, MOPS, TES, HEPES, DIPSO, MOBS, TAPSO, Tris-HCl, HEPPSO, POPSO, TEA, EPPS , trimethylglycine, Gly-Gly, N-dihydroxyethylglycine, HEPBS, TAPS, AMPD, TABS, AMPSO, CHES, CAPSO, APS, CHAPS, CABS, phosphate and histidine, or a combination thereof .

Without being bound by any theory, it is believed that the use of a buffer may help stabilize the composition at physiological pH.

The concentration of the buffer salt in the aqueous pharmaceutical formulation may range from 1 mM to 1 M, preferably from 1 mM to 100 mM, preferably from 5 mM to 50 mM, preferably from 5 mM to 20 mM.

Aqueous pharmaceutical formulations may also include counterions and salts, such as sodium counterions, chloride ions, or NaCl dissolved in the solution.

Aqueous pharmaceutical formulations may also include other active agents, such as other therapeutic or prophylactic agents.

Optionally, the aqueous pharmaceutical formulation according to the invention is essentially free of meglumine.

Optionally, the aqueous pharmaceutical formulation consists of:

water;

Optional salts, such as buffer salts or dissolved NaCl;

Compounds of formula I,

and

Compounds of formula II, Formula II

where each ^R1 is independently selected from -H or group;

wherein R ² is absent or is C _1-4 alkyl;

Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and

R ³ is independently selected from -H or C _1-4 alkyl;

wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2; and

The pH of the pharmaceutical preparation is between pH 7 and pH 8.

Aqueous pharmaceutical composition

Preferably, the aqueous pharmaceutical preparation of the present invention can be obtained by a method including:

Step 1: Prepare a mixture including water, a compound of formula I and a compound of formula II, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2;

Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8.

Preferably, step 1 is performed at a pH of 9 or higher. The pH can be set using any pharmaceutically acceptable salt, such as sodium hydroxide.

Preferably, step 1 is performed at a pH of 10 or higher, preferably at a pH of 10.5 or higher, preferably at a pH of 10.7 or higher, preferably at a pH of 11 or higher.

Any pharmaceutically acceptable acid can be used to lower the pH in step 2, such as 1-hydroxy-2-naphthoic acid, 2,2-dichloroacetic acid, 2-hydroxyethanesulfonic acid, 2-oxoglutaric acid, 4-acetylaminobenzoic acid, 4-aminosalicylic acid, acetic acid, adipic acid, ascorbic acid (L), aspartic acid (L), benzenesulfonic acid, benzoic acid, camphoric acid (+), camphor-10 -Sulfonic acid (+), caproic acid (capric acid), hexanoic acid (caproic acid), caprylic acid (octanoic acid), carbonic acid, cinnamic acid, citric acid, cyclic acid, dodecyl sulfate, ethane-1,2 -Disulfonic acid, ethanesulfonic acid, formic acid, fumaric acid, mucic acid, gentisic acid, glucoheptonic acid (D), gluconic acid (D), glucuronic acid (D), glutamic acid, glutaric acid , glycerophosphate, glycolic acid, hippuric acid, hydrobromic acid, hydrochloric acid, isobutyric acid, lactic acid (DL), lactouronic acid, lauric acid, maleic acid, malic acid (-L), malonic acid, mandelic acid ( DL), methanesulfonic acid, naphthalene-1,5-disulfonic acid, naphthalene-2-sulfonic acid, nicotinic acid, nitric acid, oleic acid, oxalic acid, palmitic acid, parapeptic acid, phosphoric acid, propionic acid, pyroglutamic acid ( -L), salicylic acid, sebacic acid, stearic acid, succinic acid, sulfuric acid, tartaric acid (+L), thiocyanic acid, toluenesulfonic acid (p) and/or undecenoic acid.

Within the meaning of the present invention, an aqueous pharmaceutical preparation may also mean a reconstituted liquid form obtainable by reconstituting a dry pharmaceutical preparation according to the invention (as described below) in a pharmaceutically acceptable grade of water, buffer solution or salt. Pharmaceutical preparations.

dry form

The invention also relates to dry pharmaceutical preparations obtainable by drying the aqueous pharmaceutical preparations according to the invention.

Without wishing to be bound by any theory, it is believed that by drying the aqueous formulation according to the present invention, it is possible to increase the stability of the compound (QTX125) and ensure that the pH of the liquid pharmaceutical composition is between pH 7 and pH 8 upon reconstitution.

The dry pharmaceutical composition according to the present invention includes a compound of formula I and a compound of formula II in a molar ratio of 1:50 to 1:2.

The drying step can be accomplished by any drying method known to those skilled in the art, such as freeze drying or spray drying. Preferably, the drying step is performed by lyophilizing the aqueous pharmaceutical formulation according to the invention.

Preferably, the dry pharmaceutical formulation can be obtained by methods including:

Step 1: Prepare a mixture including water, a compound of formula I and a compound of formula II, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2;

Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8;

Step 3: Dry the solution.

Preferably, step 1 is carried out at a pH of 9 or higher, preferably at a pH of 10 or higher, preferably at a pH of 10.5 or higher, preferably at a pH of 10.7 or higher, preferably at a pH of 11 or higher.

Without wishing to be bound by any theory, it is believed that the compound of Formula I is incorporated into the cavity of the compound of Formula II (beta-cyclodextrin) by dissolving the compound of Formula I at high pH. The compound of formula I remains bound in the cavity during steps 2 and 3, which means that the compound of formula I is soluble in aqueous solutions at physiological pH when the solution is reconstituted.

Dry pharmaceutical formulations can be reconstituted into aqueous pharmaceutical formulations according to the invention by reconstituting the formulation in pharmaceutically acceptable grade water, buffered solutions or saline solutions to obtain aqueous pharmaceutical compositions.

Alternatively, the dry pharmaceutical composition may be obtained by methods including:

Step 1: Prepare a mixture including water, a compound of formula I and a compound of formula II, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2;

Step 2: Dry the solution.

In this case, the reconstitution step is preferably performed with an acidic solution to ensure that the solution obtained is at a pH suitable for injection.

Preparation of the composition

The present invention also relates to a method for preparing an aqueous pharmaceutical preparation according to the present invention, wherein the method includes the following steps:

Step 1: Preparing a mixture including water, a compound of formula I and a compound of formula II;

Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8.

Preferably, step 1 is carried out at a pH of 9 or higher, preferably a pH of 10 or higher, preferably a pH of 10.5 or higher, preferably a pH of 10.7 or higher, preferably a pH of 11 or higher.

Preferably, the concentration of the compound of formula II in step 1 is from 10 mg/mL to 2000 mg/mL, more preferably from 50 mg/mL to 1500 mg/mL, and most preferably from 100 mg/mL to 1000 mg/mL.

Preferably, step 1 includes:

Step 1a: Preparing a mixture including water and a compound of formula II;

Step 1b: Add chlorine to the above mixture to ensure that its pH is 9 or higher, preferably pH is 10 or higher; preferably pH is 11 or higher;

Step 1c: Add the compound of formula I.

Preferably, during step 1, the mixture is stirred using a stirring device until all compounds of formula I are dissolved. The stirring device used is not particularly limited, and suitable stirring devices may include a vortex mixer, a magnetic stirrer, a spiral mixer or a paddle stirrer.

Preferably, the mixture in step 1 is stirred for at least 40 minutes. Without wishing to be bound by any theory, it is believed that this is the time necessary to ensure that the QTX125 molecules enter the cyclodextrin cavity and disrupt any intramolecular non-covalent interactions.

Optionally, when the method includes steps 1a, 1b and 1c, the mixture may be stirred for at least 20 minutes in step 1a, at least 15 minutes in step 1b, and at least 40 minutes in step 1c.

Preferably, step 2 also includes diluting the mixture with a diluent, such as water.

Preferably, the mixture is filtered after step 2, for example through a 0.45 µm or 0.2 µm filter.

The method may also involve a method for producing a dry pharmaceutical preparation, comprising the following steps:

Step 1: Preparing a mixture including water, a compound of formula I and a compound of formula II;

Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8;

Step 3: Dry the above mixture.

The preferred aspects related to steps 1 and 2 described for the method for the preparation of aqueous pharmaceutical preparations according to the invention also apply to the method for the production of dry pharmaceutical preparations.

Preferably, the drying in step 3 is performed by freeze-drying the above pharmaceutical formulation.

Optionally, the method may further comprise step 4 of storing the dry pharmaceutical formulation at room temperature for a period of at least three months.

The present invention also relates to aqueous pharmaceutical preparations or dry pharmaceutical preparations obtainable by the above-mentioned methods.

Medical uses, treatments

In another aspect, the invention relates to the use of an aqueous pharmaceutical formulation or a dry pharmaceutical formulation according to the invention for the preparation of a medicament.

Preferably, the present invention relates to the use of an aqueous pharmaceutical preparation or a dry pharmaceutical preparation according to the invention for the preparation of a medicament for the treatment of cancer.

Alternatively, the present invention relates to the use of an aqueous pharmaceutical preparation or a dry pharmaceutical preparation according to the invention for the preparation of a medicament for the treatment of autoimmune diseases.

In another aspect, the invention relates to the use of an aqueous pharmaceutical formulation according to the invention or a liquid pharmaceutical formulation obtained by reconstituting a dry pharmaceutical formulation as a medicament.

Preferably, the present invention relates to the use of an aqueous pharmaceutical preparation according to the invention or a liquid pharmaceutical preparation obtained by reconstituting a dry pharmaceutical preparation for the treatment of cancer.

Alternatively, the present invention relates to the use of an aqueous pharmaceutical formulation according to the invention or a liquid pharmaceutical formulation obtained by reconstituting a dry pharmaceutical formulation for the treatment of autoimmune diseases.

In another aspect, the invention relates to a method of treatment comprising administering to a patient in need of such treatment an aqueous pharmaceutical formulation according to the invention or a liquid pharmaceutical formulation obtained by reconstituting a dry pharmaceutical formulation.

Preferably, the present invention relates to a method of treating cancer, comprising administering to a patient in need of such treatment an aqueous pharmaceutical formulation according to the invention or a liquid pharmaceutical formulation obtained by reconstituting a dry pharmaceutical formulation.

Alternatively, the present invention relates to a method of treating an autoimmune disease, comprising administering to a patient in need of such treatment an aqueous pharmaceutical formulation according to the invention or a liquid pharmaceutical formulation obtained by reconstituting a dry pharmaceutical formulation.

Preferably, the cancer is selected from breast cancer, chronic myeloid (or myeloid) leukemia (CML), colorectal cancer, lymphoma (such as non-Hodgkin lymphoma), fibrosarcoma, gastric cancer, glioblastoma, Kidney cancer, liver cancer, lung cancer, melanoma, nasopharyngeal cancer, oral cancer, multiple myeloma in situ, osteosarcoma, ovarian cancer, pancreatic cancer and prostate cancer.

Preferably, the autoimmune disease is selected from the group consisting of autoimmune hepatitis; inflammatory demyelinating diseases of the central nervous system; systemic lupus erythematosus; acute anterior uveitis; Sjaeger's syndrome; rheumatoid arthritis ; Type 1 diabetes; Graves' disease; and inflammatory bowel disease.

Inflammatory demyelinating diseases of the central nervous system are diseases in which the myelin supporting cells of the central nervous system, such as oligodendrocytes and/or the myelin layer, are destroyed. Demyelination causes a disruption in nerve signals between the brain and the rest of the body, ultimately leading to a range of signs and symptoms, including physical, mental, and sometimes psychiatric problems.

Specific non-limiting examples of inflammatory demyelinating diseases are multiple sclerosis (MS), including relapsing MS, progressive MS, opto-spinal MS; neuromyelitis optica; acute disseminated encephalomyelitis; acute hemorrhage leukoencephalitis; Balo concentric sclerosis; Heard's disease; Marlborough MS; neoplastic MS; solitary sclerosis; optic neuritis; transverse myelitis; Susak syndrome; leukoaraiosis; myocardial infarction Painful encephalomyelitis; Guillain-Balinese syndrome; progressive inflammatory neuropathies; leukodystrophies, including adrenoleukodystrophy and adrenomedullary neuropathy. Preferably, the autoimmune disease is multiple sclerosis or acute disseminated encephalomyelitis. More specifically, it is acute disseminated encephalomyelitis, or more specifically and most preferably, it is multiple sclerosis.

Preferably, the autoimmune disease is selected from autoimmune hepatitis and inflammatory demyelinating diseases of the central nervous system.

In a particularly preferred embodiment, the autoimmune disease is an inflammatory demyelinating disease of the central nervous system as described above.

In another particularly preferred embodiment, the autoimmune disease is autoimmune hepatitis.

The inventors have found that, unlike other histone deacetylase inhibitors, QTX125 advantageously shows no evidence of genotoxicity, particularly chromosomally disruptive or non-genetic. Likewise, it was unexpectedly observed that QTX125 has improved pharmacokinetic properties compared to other histone deacetylase inhibitors, in particular a higher half-life and volume of distribution.

Apply

Preferably, the aqueous pharmaceutical formulation according to the invention (or the liquid pharmaceutical formulation obtained by reconstituting the dry pharmaceutical formulation according to the invention) is administered by injection. Pharmaceutical formulations can be administered by infusion (continuous) or bolus (intermittent) administration.

The method of administration by injection may be, for example, subcutaneous, intradermal, intramuscular, intravenous, intraarterial, intracardiac, intrathecal, intraspinal, intracapsular, subcapsular, intraorbital, intraperitoneal, intratracheal, subcutaneous, articular Intrasternal, subarachnoid, and intrasternal injections.

Preferably, administration is by intravenous infusion or intravenous injection (bolus administration). More preferably, administration is by intravenous infusion.

Subject/Dose

The subject of administration can be any animal. Preferably, the subject is a mammal, such as a rat, mouse, feline, canine, equine, porcine, ovine, bovine, primate or human. Preferably, the subject is a human patient.

Generally speaking, the effective amount of a compound of formula I to be administered will depend on a number of factors, such as the severity of the disease being treated and the body weight of the subject. The active compound is usually administered one or more times per day, for example 1, 2, 3 or 4 times per day, with typical total daily doses in the range of 0.01 to 1000 mg/kg/day.

Preferably, the compound of formula I is administered to human patients at a dose of 0.5 to 50 mg/kg, preferably 0.5 to 30 mg/kg, preferably 1 to 20 mg/kg, more preferably 5 to 10 mg/kg.

Preferably, the compound of formula I is administered to human patients at a dose of 25 mg to 4500 mg, preferably 50 mg to 3000 mg, preferably 250 mg to 1500 mg per day.

The compounds of the invention can be used with at least one other drug to provide combination therapy. Such other drug or drugs may be part of the same composition, or may be provided as separate compositions and may be administered at the same time or at different times.

Test kit

Another aspect of the invention relates to a kit comprising a dry (preferably freeze-dried) pharmaceutical formulation according to the invention and a pharmaceutically acceptable grade of water, buffer solution or saline solution for reconstituting the dosage form. Preferably, the pharmaceutical composition is provided in a suitable container and/or has suitable packaging.

Kits may also include one or more delivery systems, such as syringes and needles, for delivering or administering the pharmaceutical compositions provided therein. The kit may also include instructions for use (eg, instructions for treating a subject).

Preferably, the kit also includes instructions for use, eg written instructions on how to administer the composition (eg injection regimen). Most preferably, the kit includes written instructions on how to prepare the appropriate pharmaceutical formulation from the dry pharmaceutical formulation (eg, how to reconstitute the formulation) and how to subsequently administer the reconstituted pharmaceutical formulation.

As those skilled in the art will appreciate, features and preferred embodiments of one aspect of the invention will also apply to other aspects of the invention.

Preferred embodiment

Particularly preferred embodiments include:

An aqueous pharmaceutical preparation for injection, including:

A compound of formula I dissolved in a concentration of 9 mg/mL or higher; and

Sulfobutyl ether-β-cyclodextrin

The molar ratio of the compound of formula I to sulfobutyl ether-β-cyclodextrin is 1:15 to 1:2.3.

Preferably, the ratio of compound of formula I to sulfobutyl ether-β-cyclodextrin is from 1:4.5 to 1:2.3.

Preferably, the aqueous pharmaceutical composition can be obtained by a method including:

Step 1: Prepare a mixture including water, a compound of formula I and sulfobutyl ether-β-cyclodextrin, wherein the molar ratio of the compound of formula I to sulfobutyl ether-β-cyclodextrin is 1:15-1 :2.3, and optionally wherein the pH of the mixture is 9 or higher;

Step 2: Add acid to lower the pH of the mix to a value between pH 7 and pH 8.

In a further preferred embodiment, the present invention relates to a dry pharmaceutical preparation, which can be obtained by freeze-drying the above-mentioned aqueous pharmaceutical preparation, which includes:

A compound of formula I dissolved in a concentration of 9 mg/mL or higher; and

Compounds of formula II;

wherein the molar ratio of the compound of formula I to the compound of formula II is 1:15 to 1:2.3,

Dry pharmaceutical preparations can be obtained by the following methods:

Step 1: Prepare a mixture including water, a compound of formula I and a compound of formula II, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2.3, and wherein the pH of the mixture is 9 or more , preferably 10.5 or more;

Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8;

Step 3: Freeze-dry the above pharmaceutical preparation.

Preferably, the compound of formula II is sulfobutyl ether-β-cyclodextrin (SBβCD).

Preferably, the pH of the mixture in step 1 is 10.7 or higher.

In a further preferred embodiment, the present invention relates to a kit comprising a freeze-dried pharmaceutical formulation as described above and a pharmaceutically acceptable grade of water, buffer solution or saline solution for reconstituting the dosage form.

In a further preferred embodiment, the invention relates to the use of a pharmaceutical preparation or a freeze-dried preparation as described above for the preparation of a medicament.

Preferably, the medicament is used to treat cancer or autoimmune diseases.

More preferably, the medicament is used to treat cancer.

Example

In order to better understand the nature of the invention, some illustrative examples will now be described.

The scope of the present invention is not limited to the examples provided below. These examples merely demonstrate the effectiveness of the invention.

The experimental results in WO 2011/039353 are incorporated herein by reference.

Experiment 1 – Formulation Development

This example demonstrates how the solubility of various aqueous formulations including compounds of formula I varies depending on the formulation of the aqueous solution. Formula I (3-(3-furyl)-N-{4-[(hydroxylamino)carbonyl]benzyl}-5-(4-hydroxyphenyl)-1H-pyrrole-2-methamide (QTX125))

Excipients

Parenteral grade excipients were used in the formulation development in Example 1. Excipient details are listed in Table 1 below. Table 1: Excipient details Element manufacturer Batchno./Lotno. sodium hydroxide Avantor 0000130791 hydrochloric acid Merck KGaA CG8C680514 Glacial acetic acid Avantor 0000152649 meglumine Spectrum 3GD0046 sodium carbonate Merck KGaA DA6D660033 L-arginine Shanghai Kyowa 17052625 Phosphate buffer solution, pH 7.20 Sigma life sciences SLB57363 dimethylacetamide Panreac 869004 propylene glycol Croda 00012111510 PVP K30/Kollidon 30LP BASF 12355816K0 Hydroxypropyl-β-cyclodextrin (average molecular weight: 1541.5 g/mol) Shandong binzhouzhiyuan biotechnology 20180502 ethanol Panreac 827369 L-arginine Shanghai Kyowa 17052625 meglumine Spectrum 3GD0046 sodium chloride Merck KGaA K48658924 705 Sulfobutyl ether-β-cyclodextrin (Captisol) (average molecular weight: 2161.7 g/mol) Cydex Pharmaceuticals NC-04A-160149 glycerin Cargill 06-03-30012018-013 Glacial acetic acid JT Baker (Avantor) 0000152649 Soybean oil Croda 0000993635 Fat emulsion 20% v/v emulsion Fresenius kabi 8011007 PEG 400 BASF 96402588QO Water for Injection IH NA

The 0.2µm screening program is the PDVF screening program produced by Merck;

The 0.45µm screening program is a nylon needle screening program produced by Merck.

Solubility test procedure

The solubility test in this application is performed as follows:

Weigh the required amount of excipient into the vial.

Add diluent/co-solvent according to the composition of the respective test.

Vortex contents until dissolved.

Weigh and add the required amount of QTX125 to the above mixture and vortex for 10–15 minutes.

Use diluent to make up the volume of the formulation.

Mix the above solution using a bottle rotation device at room temperature for 24 hours.

Abbreviation

The following abbreviations are used in Example 1. WFI Water for Injection IH internal API QTX125(3-(3-furyl)-N-{4-[(hydroxylamino)carbonyl]benzyl}-5-(4-hydroxyphenyl)-1-pyrrole-2-methamide) IV intravenously DMA dimethylacetamide PBS Phosphate buffer solution Qs Quantity replenishment HPβCD Hydroxypropyl-β-cyclodextrin SBβCD Sulfobutyl ether-β-cyclodextrin HCl hydrochloric acid NaOH sodium hydroxide

Example 1.1 - pH adjustment

In this example, the effect of pH adjustment was examined. The results are given in Table 2. Table 2: Formulation composition for pH adjustment test Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T1 50 mgAPI+200 µL 2.0 N sodium hydroxide+4.80 mL phosphate buffered saline (pH 7.20) 10 12.34 NA Precipitate observed - not analyzed T2 50mg API+2.50 mL phosphate buffered saline (pH 7.20)+7.50 mL 0.5M tromethamine 5 10.70 NA Light brown suspension—not analyzed T3 1% API+2% L-Arginine+97% WFI 10 10.52 0.60 White to light yellow turbid suspension T4 1% API+5% meglumine+94% WFI 10 10.93 2.33 White to light yellow turbid suspension T5 50 mg API+5.0 mL 0.1M tromethamine 10 10.25 0.07 white turbid suspension T6 50 mg API+5.0 mL WFI+2.80 ml 0.1N NaOH 6.41 11.03 3.83 clear brown solution T7 1% API + 200 µl 1N NaOH + make up volume with WFI 10 11.79 6.02 light brown clear solution T8 1% API+10% meglumine+10% PVP K30+300 µl 1N NaOH+make up volume with WFI 10 11.75 9.03 Light yellow clear solution

According to this data, QTX125 was observed to form a turbid suspension. All alkalizing agents used in the study achieved pH values above 10. This does not apply to parenteral administration (without being bound by any theory, it is believed that if the pH is too high it can cause pain during administration). Therefore, pH adjustment methods alone will not help solubilize QTX125 within the physiological pH range.

Example 1.2 - Co-solvent

In this example, the effect of cosolvents on the solubility of QTX125 was examined. The results are given in Table 3. Table 3: Formulation composition for co-solvent testing Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T9 1% API+10% dimethylacetamide+30% propylene glycol+59% phosphate buffered saline (pH 7.20) 10 8.34 0.38 Light yellow turbid suspension T10 1% API+10% dimethylacetamide+10% PEG 400+79% phosphate buffered saline (pH 7.20) 10 7.37 0.15 Light yellow turbid suspension T11 1% API+10% dimethylacetamide+20% ethanol+69% phosphate buffered saline (pH 7.20) 10 8.43 0.36 Dissolved in a mixture of DMA and ethanol, it becomes a turbid suspension after adding PBS T12 1% API+30% propylene glycol+10% PEG 400+59% phosphate buffered saline (pH 7.20) 10 7.44 0.21 Light yellow turbid suspension T13 1% API+30% propylene glycol+20% ethanol+49% phosphate buffered saline (pH 7.20) 10 8.44 0.98 Light yellow turbid suspension T14 1% API+10% PEG 400+20% ethanol+69% phosphate buffered saline (pH 7.20) 10 7.60 0.20 Light yellow turbid suspension T15 1% API+10% dimethylacetamide+89% phosphate buffered saline (pH 7.20) 10 8.02 0.01 Dissolved in DMA, turns into a turbid suspension after adding PBS T16 1% API+10% dimethylacetamide+20% ethanol+69% 0.9% NaCl solution 10 6.75 0.63 A yellowish suspension and precipitate were observed T17 1% API+10% dimethylacetamide+20% ethanol+69% WFI 10 6.94 0.32 Light yellow turbid suspension T18 1% API+10% dimethylacetamide+20% ethanol+0.5% PVP K30+68.5% WFI 10 5.05 2.91 A small amount of precipitation was observed T19 1% API+30% propylene glycol+20% ethanol+49% WFI 10 7.21 2.03 White to light yellow turbid suspension T20 1% API+30% propylene glycol+20% ethanol+49% 0.9% NaCl solution 10 7.07 0.75 White to light yellow turbid suspension T21 10 mg API+1 mL soybean oil 10 NA NA API remains undissolved T22 25mg API + 1.4ml N-methylpyrrolidone + use WFI to make up to 10mL 2.5 4.86 0.26 Clear solution with dispersed particles

QTX125 remained undissolved or formed a turbid suspension in all trials using different solvent compositions at their maximum allowed doses. The use of co-solvent methods is not sufficient to dissolve QTX125.

Example 1.3-Cyclodextrin

In this example, the effect of using a formulation including cyclodextrin was examined. The results are given in Table 4. Table 4: Formulation composition for cyclodextrin testing Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T23 1% API+5% HPβCD+94% WFI 10 7.16 0.14 White to light yellow turbid suspension

HPβCD formed a turbid suspension with a solubility of 0.14 mg/mL after 24 h. It was concluded that HPβCD alone did not contribute to the solubilization of QTX125.

Example 1.4 - Cosolvents and Microemulsions

In this example, the effect of using cosolvents and microemulsions was examined. The results are given in Table 5. Table 5: Formulation composition of trials using co-solvent and fat emulsion 20% v/v emulsion Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T24 0.5% API+5% DMA+10% ethanol+84.5% fat emulsion 20% v/v emulsion 5 7.26 4.68 Milky white lotion T25 1% API+10% DMA+0.5 mL of benzyl alcohol+fat emulsion 20% v/v emulsion 10 6.97 7.32 White turbid solution with undissolved particles T26 1% API+20% ethanol+79% fat emulsion 20% v/v emulsion 10 7.16 9.85 White turbid solution with undissolved particles T27 1% API+10% DMA+10% ethanol+79% fat emulsion 20% v/v emulsion 10 7.14 6.07 White turbid solution with undissolved particles T28 0.5% API+40% tert-butyl alcohol+59.5% fat emulsion 20% v/v emulsion 5 6.30 4.85 Light yellow turbid solution T29 1% API+5% DMA+10% ethanol+2.25% glycerin+1.2% egg lecithin+WFI 10 5.39 0.22 Pale white turbid liquid with particles at the bottom of the vial T30 1% API+5% DMA+10% ethanol+2.25% glycerol+WFI 10 4.67 0.05 Pale white turbid liquid with particles at the bottom of the vial T31 0.5% API + 5% dimethyl acetamide + 10% ethanol – clear the solution after stirring and add the solution to 0.5 mL of Lipid Emulsion 20% v/v emulsion with stirring, followed by Lipid Emulsion 20% v/v Lotion increases volume to 1mL 5 5.13 1.46 Initially the API solution is miscible with the lipid emulsion 20% v/v emulsion. After stirring for 15 minutes, the emulsion appeared broken and oil droplets were visible on the top of the emulsion. After 24 hours - the lotion breaks down. Oil droplets were observed.

It was observed that the pH value of the drug solution was within the required range and the solubility obtained was as high as 9.85 mg/mL. However, the solution was unstable and the emulsion was observed to break down after 24 hours, which means that it is not suitable for the development of pharmaceutical formulations, where it is important that QTX125 remains in solution during long-term storage. To check the reproducibility of T24, trial T31 was conducted using the same concentrations of drug and excipients and following the same preparation procedure as T24. However, it was noted that the results varied from trial to trial and that it was not possible to obtain the same level of solubility reproducibly using this method. Therefore, it was concluded that this approach is not suitable for the development of IV formulations of QTX125.

Example 1.5 - Alkalizing Agents and Cosolvents

In this example, the effect of using alkalizing agents and co-solvents was examined. The results are given in Table 6. Table 6: Formulation composition for alkalinizing agent and co-solvent tests Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T32 1% API+5% dimethylacetamide+10% ethanol+0.03% WFI+83.9% 1.0N meglumine solution 10 10.52 8.06 Light yellow clear solution T33 1% API+5% dimethylacetamide+10% ethanol+84% 0.1N meglumine solution 10 10.02 3.25 Turbid solution T34 0.5% API+5% dimethylacetamide+10% ethanol+0.02% 1.0N meglumine solution+0.02% 0.1N HCl+83.9% WFI 5 9.71 0.40 Turbid solution

Example 1.6 - pH adjustment and cyclodextrin

In this example, the effects of using pH adjustment and cyclodextrin were examined. The results are given in Table 7. Table 7: Formulation composition for pH adjustment and cyclodextrin testing Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T35 1% API+20% HPβCD+300µl 1.0N NaOH+make up the volume with water for injection (WFI) 10mg 12.14 9.80 Light yellow clear solution T36 1% API+20% HPβCD+300µl 1.0N NaOH+make up the volume with 0.9% sodium chloride solution 10mg 11.94 10.61 Light yellow clear solution T37 1% API+20% HPβCD+10% meglumine+69% WFI 10mg 10.72 10.29 Light yellow clear solution T38 1% API+20% HPβCD+10% meglumine+69% 0.9% sodium chloride solution 10mg 10.71 9.63 Light yellow clear solution T39 1% API+3.3% HPβCD+95.7% 1.0N meglumine solution (the molar ratio of API to HPβCD is 1:1) 10mg 11.05 8.32 Clear yellowish solution T40 1% API+6.6% HPβCD+92.4% 1.0N meglumine solution (the molar ratio of API to HPβCD is 1:2) 10mg 11.01 9.06 Clear yellowish solution T41 1% API+3.5% SBβCD+95.5% 1.0N meglumine solution (the molar ratio of API to SBβCD is 1:1) 10mg 11.27 9.31 Clear yellowish solution T42 1% API+17.4% SBβCD+81.6% 1.0N meglumine solution (the molar ratio of API to SBβCD is 1:5) 10mg 11.27 9.53 Clear yellowish solution T43 1%+17.4% SBβCD+0.5% PVP K30+81.1% 1.0N meglumine solution 10mg 11.26 8.19 Clear yellowish solution T44 1% API+6.6% HPβCD+30% 1.0N meglumine solution+62.4% WFI 10mg 10.62 8.87 Turbid suspension T45 1% API+6.6% HPβCD +0.5% PVP K30+20% 1.0N meglumine solution+71.9% WFI 10mg 10.59 6.56 Turbid suspension T46 1% API+17.4% SBβCD+20.0% 1.0N meglumine solution+61.6% WFI 10mg 10.79 9.75 Clear yellowish solution T47 1% API+17.4% SBβCD+0.5% PVP K30+20.0% 1.0N meglumine solution+61.1% WFI 10mg 10.75 9.67 Clear yellowish solution T48 1% API+6.59% HPβCD+0.01% WFI+use 1.0 N meglumine solution to final volume 10mg 10.92 4.94 Light yellow clear solution T49 1% API+17.38% SBβCD+0.02% WFI+0.04% 0.1N acetic acid+use 1.0N meglumine solution to final volume 10mg 10.87 4.32 Light yellow clear solution T50 0.5% API + 6.59% HPβCD + 0.01% WFI + use 1.0N meglumine solution to final volume 5mg 11.08 2.60 Light yellow clear solution T51 0.5% API + 0.03% WFI + 0.052% acetic acid, 0.1N + 17.38% SBβCD + use 1.0 N meglumine solution to final volume 5mg 9.99 2.38 Light yellow with dispersed particles T52 API+17.38% SBβCD +0.02% WFI+0.19% acetic acid + use 1.0N meglumine solution to final volume (strength: 4mg/mL) 4mg 9.81 2.06 Light yellow with dispersed particles T53 Step 1: Mix 0.45mL WFI+8mg API +0.05mL 2.0N NaOH+66mg HPβCD Step 2: Adjust pH with 0.5 mL of 0.1N HCl 8mg 9.22 3.37 The solution is turbid with scattered particles T54 Step 1: Mix 0.45mL WFI+8mg API+0.05mL 2.0N NaOH+66mg SBβCD Step 2: Adjust pH with 0.5mL 0.1N HCl 8mg 8.91 2.61 slightly turbid solution T55 Step 1: Mix 0.45mL WFI+8mg API+0.05 mL 2N NaOH+66mg HPβCD Step 2: Adjust pH with 0.56mL 0.1N HCl 8mg 8.64 2.53 Pink turbid liquid with particles at the bottom of the vial T56 Step 1: Mix 0.45 mL WFI+8mg API+0.05 mL 2N NaOH+66mg SBβCD Step 2: Adjust pH with 0.57mL of 0.1N HCl 8mg 7.48 1.29 Light pink turbid liquid with particles at the bottom of the vial ^T57b Step 1: Mix 0.45ml WFI+8mg API +0.05ml 2N NaOH+174 mg SBβCD Step 2: Adjust the pH with 0.66 mL 0.1N HCl, reduce the pH to 2.20, add 3 drops of 0.1N NaOH solution - pH2.30 - bring to 1.50 mL of this solution was transferred to another vial and analyzed for solubility. 8mg 8.75 4.16 Very slightly turbid solution T58 Test 55 with pH adjusted to 2.30 8mg 2.30 4.75 Very slightly turbid solution T59 Step 1: Mix 0.45ml WFI+4mg API+0.05ml 2N NaOH+55.6mg SBβCD Step 2: Adjust pH with 0.72 mL 0.1N HCl 1. Unfiltered 2. Filtered through 0.45µm nylon needle screener 4mg 8.14 1. 3.75 2. 3.63 ^a Light yellow clear solution T60 Step 1: Mix 0.2ml WFI+10mg API +0.05ml 2N NaOH+200mg SBβCD Step 2: Adjust pH with 0.72 mL 0.1N HCl+0.08 mL 0.1N NaOH 1. Unfiltered 2. Filtered through 0.45µm nylon needle screening program 8mg 7.36 1. 6.95 2. 6.96 Light yellow clear solution T61 Step 1: Mix 0.15 ml WFI+10 mg API +0.05ml 2N NaOH +200 mg SBβCD Step 2: Adjust pH with 0.227 mL 0.5N HCl+5µL 0.1N HCl+0.026 mL 1.0N NaOH 1. Unfiltered 2. Pass 0.45 µm nylon needle screening program filtration 10mg 7.61 1. 8.91 2. 8.85 Light yellow clear solution T62 Step 1: Mix 0.15 ml WFI+10 mg API +0.06ml 1N NaOH +200 mg SBβCD Step 2: Adjust pH with 0.2 mL 0.25N HCl+0.02 mL 0.1N HCl+5µL 0.1N NaOH 1. Unfiltered 2. Pass 0.45 µm nylon needle screening program filtration 10 mg 7.40 1. 9.48 2. 9.33 Light yellow clear solution T63 Step 1: Mix 0.20 ml WFI+200mg SBβCD+10mg API+0.06 ml 1N NaOH Step 2: Adjust pH with 0.21mL 0.25N HCl+0.04mL 0.1N HCl 1. Unfiltered 2. Filtered through 0.45µm nylon needle screener 10mg 6.88 1. 8.01 2. 8.04 clear colorless solution T64 Step 1: Mix 0.30ml WFI+139mg SBβCD+10mg API+0.05 ml 1N NaOH, Step 2: Adjust the pH with 0.19 mL 0.25N HCl+0.01 mL 0.1N NaOH (the ratio of drug to cyclodextrin is 1:4) 10mg 7.26 9.29 Light yellow clear solution T65 Step 1: Mix 0.30 ml WFI+104mg SBβCD+10mg API+0.06ml 1N NaOH Step 2: Adjust the pH with 0.26 mL 0.25N HCl+0.018 mL 0.1N HCl (the ratio of drug to cyclodextrin is 1:3) 10mg 7.45 7.54 Light yellow solution, precipitation was observed after being left at room temperature for 24 hours. Numbers 1 and 2 in the solubility column in ^table a refer to the solubility of QTX125 in filtered and unfiltered versions of the solution. ^b In this experiment, the pH dropped to pH 2.2 and then increased to pH 2.3. The solution at pH 2.3 was then used to determine the solubility of the compound.

Example 1.7 - pH adjustment and freeze drying

In this example, the effect of using pH adjustment and freeze-drying was examined. The results are given in Table 8.

Samples were prepared as described above in the solubility test protocol section, and the solutions were then freeze-dried. The solution was then reconstituted with 10 mL of water and the solubility of QTX125 was determined. Table 8: Formulation composition of pH adjuster and freeze-drying tests Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T66 1% API + 15.4% sodium carbonate + use WFI to 5mL – freeze dry 10 11.53 3.54 dark yellow clear solution T67 1% API + 15.4% sodium carbonate + 1.50ml 1.0N sodium hydroxide + use WFI to make up to 5mL – freeze drying 10 12.91 4.17 dark yellow clear solution T68 1% API + 1ml 1.0N sodium hydroxide solution + use WFI to make up to 10mL – divide into two equal parts, 1. Keep one part as it is, and 2. Add 0.5mL 0.5N acetic acid to the other part, freeze-dry 10 Part 1 I:12.67 and II: ^11.82a 6.91 1. Clarify the solution Part 2 I:11.30 and II: ^10.49a 7.57 clear solution ^a I refers to the pH measured at time 0; II refers to the pH measured 24 hours later.

Example 1.8 - pH adjustment, cosolvents and cyclodextrins

In this example, the effects of using pH adjustment, co-solvents and cyclodextrins were examined. The results are given in Table 9. Table 9: Formulation composition of pH adjuster, co-solvent and cyclodextrin tests Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T69 1% API+6.6% HPβCD+5% dimethylacetamide+0.5% PVP K30+10% ethanol+76.9% 1.0N meglumine solution 10mg 10.88 8.79 Clear yellowish solution T70 1% API+17.4% SBβCD+5% dimethylacetamide+0.5% PVP K30+10% ethanol+66.1% 1.0N meglumine solution 10mg 11.21 9.27 Clear yellowish solution

Example 1.9 - Cosolvents and cyclodextrins

In this example, the effect of using cosolvents and cyclodextrins was examined. The results are given in Table 10. Table 10: Formulation composition for cosolvent and cyclodextrin tests Test number Preparation composition Strength of initially added QTX125 (mg/mL) pH Solubility(mg/mL) Appearance T71 1% API+6.6% HPβCD+5% dimethylacetamide+10% ethanol+77.4% WFI 10mg 8.51 1.00 Off-white turbid suspension T72 1% API+17.4% SBβCD+5% dimethylacetamide+10% ethanol+66.6% WFI 10mg 4.98 0.93 Off-white turbid suspension T73 1% API+6.6% HPβCD+5% dimethylacetamide+0.5% PVP K30+10% ethanol+76.9% WFI 10mg 6.20 0.39 Off-white turbid suspension T74 1% API+17.4% SBβCD+5% dimethylacetamide+0.5% PVP K30+10% ethanol+66.1% WFI 10mg 5.41 0.95 Off-white turbid suspension

Experiment 2 – Stability and freeze-drying studies

This experiment investigated the development of a robust and stable intravenous formulation of QTX125. This study evaluates the long-term stability of such formulations.

Material

The materials used in Example 2 are provided in Table 11.

Table 11: Materials used in Example 2

HPLC measurement method

Listed below are the parameters required for the HPLC method used to determine the concentration of QTX125 in solution. Chemical/reagent name Level supplier Orthophosphate AR level Merck DMSO HPLC grade Spectrochem Methanol HPLC grade Merck Acetonitrile HPLC grade Merck Water (HPLC grade or Milli Q water) HPLC(MilliQ) Evoqua

Instruments and equipment: system Waters HPLC Alliance Pump Quaternary pump/binary pump detector UV/PDA detector

Chromatographic parameters: HPLC column Accucore C18 150mm x 4.6mm; 2.6μm (or) equivalent column (Manufacturer: Thermo) Column temperature 25±2℃ Sampler temperature 20±2℃ Detector wavelength 254nm flow rate 1.0ml/min Injection volume 5μL Execution time 55 minutes mobile phase Mobile phase A: 0.1% OPA aqueous solution Mobile phase B: acetonitrile: methanol (80:20% v/v) gradient conditions Thinner 0.1% OPA: methanol (50:50v/v)

Use the protocol above to determine the HPLC determination of the percentage of QTX125 in the following solution. The peak value of QTX125 occurs at a dwell time of approximately 24.1 minutes.

Assays (%LC) were determined by comparing the peak area of QTX125 to a calibration curve determined by running samples of known concentrations of QTX125 (standard) on HPLC.

QTX125% is calculated based on the QTX125 peak area compared to the sum of all peak areas in the chromatogram.

The percentage of impurities (%w/w) was calculated based on the area normalization method. This is based on a comparison of the relative areas of the impurity peaks (i.e. peaks with a dwell time other than 24.1 minutes) compared to the sum of the areas of all peaks in the chromatogram.

Example 2.1-1:2.7 Liquid injection formulation

Manufacturing process

The liquid injection preparation used in Example 2.1 was prepared as follows, with a batch size of 400 mL:

Add 30% v/v water based on the total volume of the formulation to the beaker;

Add the required amount of SBβCD to the beaker and stir for 20 minutes to dissolve. Check the pH of the solution.

Add 20 mL of 1.0 N NaOH (corresponding to 5% v/v NaOH) to the solution (800 mg sodium hydroxide) and stir for 15 minutes to mix. Check the pH of the solution.

Add an appropriate amount of QTX125 to the solution and stir for 40 minutes at room temperature to dissolve. Check the pH of the solution and adjust the pH to pH 7.3 using 0.25N HCl solution.

Make up the volume of the bulk solution to 100% v/v of the total batch with water for injection and stir for 15 minutes. Check the pH of the solution.

The solution was filtered through a 0.2µm PVDF membrane (Merck).

Liquid preparations with a drug to SBβCD ratio of 1:2.7 are stored at 5±3°C, 25°C 60% relative humidity (25°C/60%RH) and 40°C 75% relative humidity (40°C/75%RH) 1 The stability results after one month (1M), two months (2M) and three months (3M) are shown in Table 12.

Table 12: Stability results of 1:2.7 liquid formulation Conditions/test parameters initial 5±3℃ 25℃/60%RH 40℃/75%RH 1M 2M 3M 1M 2M 3M 1M 2M 3M describe Clear light straw color solution Clear light straw color solution Turbid straw-colored solution with dispersed particles Dark turbid straw-colored solution with dispersed particles pH 7.29 7.26 7.27 7.18 6.21 4.56 4.47 6.98 6.69 6.31 Test(%LC) 98.6 99.3 100.2 97.2 97.4 98.5 97.3 97.6 97.8 95 Related substances (%) QTX125 99.21 99.15 99.035 99.065 98.897 98.938 98.799 98.186 97.318 96.707 total impurities 0.793 0.8 0.828 0.796 1.026 0.963 1.132 1.744 2.503 3.194

Precipitation was observed in samples stored at 25°C/60%RH and 40°C/75%RH. A decrease in pH was observed in the 25°C/60% RH and 40°C/75%RH samples. Although the formulation was shown to be stable for up to 3 months at 2-8°C, it was found to be unstable at 25°C/60% RH and 40°C/75% RH, as is evident from the description of the solution, pH Decrease and total impurities increase.

Example 2.2-1:5.4 Liquid injection formulation

Samples were prepared as described in Example 2.1 above. Liquid preparations with a drug to SBβCD ratio of 1:5.4 are stored at 5±3°C, 25°C relative humidity (25°C/60%RH) and 40°C, 75% relative humidity (40°C/75%RH) 1 The stability results after one month (1M), two months (2M) and three months (3M) are shown in Table 13.

Table 13: Stability results of 1:5.4 liquid injection formulation Conditions/test parameters initial 5±3℃ 25℃/60%RH 40℃/75%RH 1M 2M 3M 1M 2M 3M 1M 2M 3M describe Clear light straw color solution Clear light straw color solution Clear light straw color solution Clear dark straw colored solution pH 7.39 7.2 7.24 7.22 7.15 7.13 7.04 6.96 6.82 6.94 Test(%LC) 98.4 98.6 98 96.6 98.4 97.5 95.7 94.1 93.9 91.1 Related substances (%) QTX125 98.973 98.927 98.886 98.936 98.569 98.312 99.065 97.345 96..098 95.592 total impurities 0.94 1.036 1.005 0.866 1.319 1.65 0.853 2.55 3.841 4.233

Example 2.3-1:2.7 Buffered liquid injection

Manufacturing process

The liquid injection preparation used in Example 2.3 was prepared as follows, with a batch size of 400 mL:

Add 30% v/v water based on the total volume of the formulation to the beaker;

Add the required amount of SBβCD to the beaker and stir for 20 minutes to dissolve. Check the pH of the solution.

Add 10 mL of 1.0 N NaOH (2.5% v/v NaOH) to the solution (400 mg sodium hydroxide) and stir for 15 minutes to mix. Check the pH of the solution.

Add an appropriate amount of QTX125 to the solution and stir for 40 minutes at room temperature to dissolve. Check the pH of the solution and adjust the pH to pH 7.3 using 0.25N HCl solution.

120 mL of 0.2N sodium dihydrogen phosphate solution (30% v/v) was added to the mixture and mixed for 15 minutes. Check the pH of the solution.

Make up the volume of the bulk solution to 100% v/v of the total batch with water for injection and stir for 15 minutes. Check the pH of the solution.

The solution was filtered through a 0.2µm PVDF membrane (Merck).

Liquid formulations with a drug to SBβCD ratio of 1:2.7 in buffer are tested at 5±3°C, 25°C and 60% relative humidity (25°C/60%RH), and 40°C and 75% relative humidity (40°C/75% The stability results after storage for 1 month (1M), 2 months (2M) and 3 months (3M) under RH) are shown in Table 14.

Table 14: Stability results of 1:2.7 buffered liquid injection

After the second and third months, lumps appeared in the samples; therefore, these samples were not tested.

Example 2.4-1:2.7 Freeze-dried products

Samples were prepared as described in Example 2.1 above, then filled into 20 mL/20 mm amber USP Type I vials, stoppered halfway with a 20 mm rubber stopper and loaded into a lyophilizer.

Use the following program for the freeze-drying process, as shown in Table 15.

Table 15: Freeze-drying process used to prepare stability test samples stage steps Temperature(℃) Time(Mins) Vacuum(Torr) bevel/hold heat treatment 1 25 10 - H 2 -5 120 - R 3 -5 60 - H 4 -40 140 - R 5 -40 360 - H extra freeze Condenser temperature: -50℃ -40 20 200 - dry 1 -40 10 200 H 2 -15 300 100 R 3 -15 900 100 H 4 -5 180 100 R 5 -5 240 100 H 6 15 180 100 R 7 15 360 100 H 8 30 240 100 R 9 30 600 100 H Afterheat 10 35 800 100 -

Stored at 5±3℃, 25℃ and 60% relative humidity (25℃/60%RH), and 40℃ and 75% relative humidity (40℃/75%RH) for 1 month (1M), 2 months After (2M) and 3 months (3M), the stability results of the reconstituted lyophilized preparation with a drug to SBβCD ratio of 1:4 are shown in Table 16.

Table 16: Stability results of 1:2.7 freeze-dried formulations Conditions/test parameters initial 5±3℃ 25℃/60%RH 40℃/75%RH 1M 2M 3M 1M 2M 3M 1M 2M 3M Description of freeze-dried cakes Off-white freeze-dried cake Reconstruction time 48sec 42 seconds ND 46sec 50sec ND 51sec 46sec ND 45sec Reconstitution solution description Clear light straw color solution Clear light straw color solution ND Clear light straw color solution Clear light straw color solution ND Clear light straw color solution Clear light straw color solution ND Clear light straw color solution Reconstitute solution pH 7.41 7.44 ND 7.51 7.45 ND 7.54 7.46 ND 7.52 water content 1.17 1.61 ND 1.77 1.27 ND 1.65 1.26 ND 1.68 Test (%LC) ^a 101.4 100.1 105.5 101.8 101.5 106.2 100.2 100.2 103.1 97.5 Related substances (%) QTX125 99.19 99.21 98.97 99.32 99.15 98.91 99.398 99.17 98.85 99.358 total impurities 0.808 0.793 0.909 0.606 0.852 0.977 0.528 0.832 0.997 0.563 ^aThe test is determined by comparison to a QTX125 standard of known weight

No changes were observed in the parameters tested. The drug product in freeze-dried cake form was stable under all three conditions (2°C-8°C, 25°C/60%RH and 40°C/75%RH) for up to 3 months.

Conclusion of Example 2

In summary, freeze-dried drug products are relatively stable relative to ready-to-use injectable forms. A summary of the observations from each of Examples 2.1-2.4 is given in Table 17 below.

Table 17: Summary of observations Example Drug to SBβCD ratio observation observe 5±3℃ 25℃/60%RH 40℃/75%RH 2.1(SF21000162) 1:2.7 (liquid injection) Stable up to 3M unstable unstable 2.4 (SF21000175) 1:2.7 (Lyo product) Stable up to 3M Stable up to 3M Stable up to 3M 2.2 (SF21000182) 1:5.4 (liquid injection) Stable up to 3M Stable up to 3M unstable 2.3(SF21000273) 1:2.7 (buffered liquid injection) Unstable precipitation observed Unstable precipitation observed Unstable precipitation observed

Example 3 - Tolerance of new formulations in Sprague Dawley rats

The purpose of this study was to evaluate the tolerance of various QTX125 formulations in mammals (male Sprague Dawley rats).

Preparations used in this example

Preparation A: 5% v/v 1N NaOH + 13.9% SBβCD (Captisol) + adjust pH to 7.2 with 0.25N HCl and/or 0.1NaOH + appropriate amount of water for injection

Preparation B: 20% PEG 400+30% propylene glycol+15% glycerin+10% Tween80+appropriate amount of 25% water for injection

The research design of this study is summarized in Table 18 below.

Table 18: Study design for testing rat tolerance to various dosage forms of different QTX125 formulations handle Number of rats Nominal dose (mg/kg) Ratio of cyclodextrin (SBβCD) and API (Mw2162g/mol) Dosage volume (mL/kg) Strength(mg/mL) Preparation A 3 15 1:35 20 0.750 3 30 1:18 1.5 3 40 1:13 2.0 3 60 1:8.9 3.0 3 80 1:6.6 4.0 3 120 1:4.4 6.0 3 200 1:2.7 10.0 Preparation B 3 30 - 1.5

Prepare the formulation on the day of dosing. The formulation was administered as an intravenous infusion to the animals of the corresponding group over 30 minutes through the femoral vein cannula using a Harvard pump.

result

Intravenous infusion of Formulation A (doses 15, 30, 40, 60, 80, 120 and 200 mg/kg) was well tolerated by male Sprague Dawley rats.

After a single intravenous infusion of Formulation B into male Sprague Dawley rats (dose: 30 mg/kg), all animals were found to die during the infusion.

These results indicate that Formulation A is better tolerated than Formulation B in male Sprague Dawley rats.

Claims (23)

An aqueous pharmaceutical preparation for injection, comprising: a compound of formula I, Formula I and compounds of formula II, Formula II where each ^R1 is independently selected from -H or The group; wherein, R ² is absent or is C _1-4 alkyl; Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and R ³ is independently selected from -H or C _1-4 alkyl; wherein, the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2; and wherein, the pH of the pharmaceutical preparation Between pH 7 and pH 8. The aqueous pharmaceutical preparation according to claim 1, wherein the compound of formula I is dissolved at a concentration of 7.5 mg/mL or higher. The aqueous pharmaceutical preparation according to claim 1 or 2, wherein the compound of formula I is dissolved at a concentration of 8 mg/mL or higher, preferably 8.5 mg/mL or higher, more preferably 9 mg/mL or higher , more preferably 9.5 mg/mL or higher, and optionally dissolved in concentrations up to a maximum concentration of 20 mg/mL. The aqueous pharmaceutical preparation according to any one of claims 1 to 3, wherein the compound of formula II is selected from β-cyclodextrin, (C _1-4 alkyl)-β-cyclodextrin, (hydroxyl -C _1-4 alkyl)-β-cyclodextrin and the group of sulfobutyl ethers of β-cyclodextrin. The aqueous pharmaceutical preparation according to any one of claims 1 to 4, wherein the compound of formula II is hydroxypropyl β-cyclodextrin or sulfobutyl ether-β cyclodextrin (SBβCD). The aqueous pharmaceutical preparation according to any one of claims 1 to 5, wherein the compound of formula II is sulfobutyl ether-β-cyclodextrin (SBβCD). The aqueous pharmaceutical preparation according to any one of claims 1 to 6, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:40 to 1:2.5, preferably 1:30 to 1:2.5 , preferably 1:25 to 1:2.5, preferably 1:20 to 1:2.5. The aqueous pharmaceutical preparation according to claim 7, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:15 to 1:2.5, preferably 1:10 to 1:2.5, preferably 1:9 to 1: 2.5, preferably 1:8 to 1:2.5, preferably 1:6 to 1:2.5, more preferably 1:4.5 to 1:2.5. The aqueous pharmaceutical preparation according to any one of the preceding claims, wherein: The compound of Formula I is dissolved at a concentration of 9 mg/mL or higher; The compound of formula II is sulfobutyl ether-β-cyclodextrin; and The molar ratio of the compound of formula I to sulfobutyl ether-β-cyclodextrin is 1:4.5 to 1:2.5. An aqueous pharmaceutical preparation for injection according to any one of the preceding claims, comprising: a compound of formula I, Formula I and compounds of formula II, Formula II where each ^R1 is independently selected from -H or The group; wherein, R ² is absent or is C _1-4 alkyl; Q is selected from the group of -H, -SO ₃ ^- , -OH, -C(O)R ³ or -C(OH)R ³ ₂ ; and R ³ is independently selected from -H or C _1-4 alkyl; wherein, the pharmaceutical preparation can be obtained by a method including the following: Step 1: Preparation of water, a compound of formula I and a compound of formula II A mixture, wherein the molar ratio of the compound of formula I to the compound of formula II is 1:50-1:2; Step 2: Adding acid to reduce the pH of the mixture to a value between pH 7 and pH 8; Optionally, wherein step 1 is performed at a pH of 9 or higher. A dry pharmaceutical preparation can be obtained by drying the aqueous pharmaceutical preparation according to any one of the preceding claims. The dry pharmaceutical preparation according to claim 11 can be obtained by freeze-drying the aqueous pharmaceutical preparation according to any one of claims 1 to 10. A kit comprising the dry pharmaceutical preparation according to claim 11 or 12 and a pharmaceutically acceptable grade of water, buffer solution or saline solution for reconstituting the dosage form. Use of the aqueous pharmaceutical preparation according to any one of claims 1 to 10 or the dry pharmaceutical preparation according to claim 11 or 12 in the preparation of medicines. Use of the aqueous pharmaceutical preparation according to any one of claims 1 to 10 or the dry pharmaceutical preparation according to claim 11 or 12 in the preparation of medicaments for the treatment of cancer or autoimmune diseases. Use of an aqueous pharmaceutical preparation according to any one of claims 1 to 10 or a reconstituted liquid pharmaceutical preparation obtainable by reconstituting a dry pharmaceutical preparation according to claim 11 or 12 as a medicament. An aqueous pharmaceutical preparation according to any one of claims 1 to 10 or a reconstituted liquid pharmaceutical preparation obtainable by reconstituting a dry pharmaceutical preparation according to claim 11 or 12 for use in the treatment of cancer or autoimmune diseases The purpose of the drug. A method of treatment comprising administering to a patient in need of such treatment an aqueous pharmaceutical formulation according to any one of claims 1 to 10 or a reconstituted dry pharmaceutical formulation obtainable according to claim 11 or 12. Structured liquid pharmaceutical preparations. The treatment method according to claim 17, wherein the method is used to treat cancer or autoimmune diseases. A method for preparing the aqueous pharmaceutical preparation according to any one of claims 1 to 10 or the dry pharmaceutical preparation according to claim 11 or 12, comprising: Step 1: Preparing a mixture including water, a compound of formula I and a compound of formula II; Step 2: Add acid to lower the pH of the mixture to a value between pH 7 and pH 8. The method according to claim 20, wherein step 1 is performed at a pH of 9 or higher. The method for preparing a dry pharmaceutical preparation according to claim 20 or 21, further comprising the step of freeze-drying the aqueous pharmaceutical preparation obtained after step 2. The method according to any one of claims 20 to 22, wherein the method further comprises the step of storing the aqueous pharmaceutical formulation or freeze-dried formulation at room temperature for a period of at least three months.